酶的固定化4.ppt

发展趋势 理性设计酶的固定化技术 In silico analysis to predict the orientation of enzymes An array of algorithms (based on the protein size and the textural properties of the support) for the rational design of immobilized derivatives Ordered mesoporous materials Tailor immobilized enzymes with high volumetric activity Fluorescence confocal microscopy Understand the diffusional restrictions and the distribution of biomolecules within the support Introduction The importance of enzyme immobilization Easy separation and reuse of the biocatalyst, makes product recovery easier and very often enhances enzyme stability For analytical, energetic and biomedical applications (e.g., biosensors, biofuel cells) Enzyme immobilization protocols Enzyme binding to a prefabricated support Enzyme entrapment or encapsulation Carrier-free cross-linking with bifunctional regents Design a robust immobilized biocatalyst Non-catalytic requirements: separation, reuse, downstream processing Catalytic function: productivity, space-time yield, productivity Irrational vs rational (by predicting the location of amino acid residues or protein domains implicated in the binding with the support) In silico analysis Hudson et al. J. Phys. Chem. B, Vol. 109, No. 41 cytochrome c and xylanase physicochemical properties of mesoporous silica surface potential of the biomolecules Weber et al. J. Mol. Catal. B-Enzym, Vol 64 They studied the adsorption of P450 enzymes on mesoporous MCM-41 and SBA-15 modeling the 3D enzyme structure performing electrostatic potential calculations predicted the pH-dependence of the P450 immobilization proposed the possible orientations of the protein on such mesoporous materials Selection of a proper support Computational Methods Applied to Covalent Binding of Enzymes to Supports Reactivity of the amino acid residues intrinsic chemical nature the microenvironment and, state of ionization LIGRe algorithm: it represents the proportion between active (e.g., deprotonated in the case of NH2 ) a